5-(substitutedphenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione analogues as anti-inflammatory agents

ABSTRACT

Compounds for treating an inflammatory disease, disorder, or condition and, particularly, to compounds that are 5-(substitutedphenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione analogues and their use as anti-inflammatory agents.

BACKGROUND

1. FIELD

The present disclosure relates to compounds having anti-inflammatoryproperties and, particularly, to anti-inflammatory compounds that are5-(substitutedphenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dioneanalogues.

2. DESCRIPTION OF THE RELATED ART

Non-steroidal anti-inflammatory drugs (NSAIDs) have been therapeuticallyused in the medication of rheumatic arthritis and in the treatment ofvarious inflammatory disorders. However, due to their gastrointestinalside effects, these have been used in limited numbers.

Thus, anti-inflammatory compounds having reduced side effects, such asreduced gastrointestinal side effects, solving the aforementionedproblems are desired.

SUMMARY

The present subject matter relates to the field of pharmaceuticals,particularly to novel5-(substitutedphenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dioneanalogues, a green synthetic process for making the same, compositionscomprising these compounds, and the use of the compounds asanti-inflammatory agents. These compounds can be made by a greensynthetic method and purified by recrystallization and columnchromatographic methods. The compounds can be obtained in good yields.Structural elucidation of the compounds is completed by spectraltechniques such as FT-IR, NMR (¹H and ¹³C), LC-MS, and elementalanalysis. These compounds show promising anti-inflammatory activitybetween millimolar to micromolar concentrations compared to standardanti-inflammatory drugs. Some of the selected lead compounds can besuccessfully taken forward to develop novel anti-inflammatory drugcandidates.

In an embodiment, the present subject matter relates to compound havingthe formula 4:

or a pharmaceutically acceptable salt, ester, stereoisomer, or solvatethereof, wherein:

-   R₁ is selected from the group consisting of hydrogen, hydroxy, and    benzyloxy;-   R₂ is selected from the group consisting of hydrogen, a C₁-C₆    straight or branched chain alkoxy, and benzyloxy;-   R₃ is selected from the group consisting of hydrogen and a C₁-C₆    straight or branched chain alkyl; and-   R₄ is selected from the group consisting of hydrogen and a C₁-C₆    straight or branched chain alkyl.

In another embodiment, the present subject matter relates to a compoundhaving the formula 4:

or a pharmaceutically acceptable salt, ester, stereoisomer, or solvatethereof, wherein:

-   R₁ is selected from the group consisting of hydrogen, hydroxy, and    benzyloxy;-   R₂ is selected from the group consisting of hydrogen, methoxy, and    benzyloxy;-   R₃ is selected from the group consisting of hydrogen and methyl; and-   R₄ is selected from the group consisting of hydrogen and methyl.

In a further embodiment, the present subject matter relates to acompound selected from the group consisting of:5-(3-(Benzyloxy)phenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione (4a);5-(4-(Benzyloxy)phenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4b);5-(3-Hydroxy-4-methoxyphenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4c);5-(3-Hydroxy-4-methoxyphenyl)-7-imino-1,3-dimethyl-7,8-dihydropyrimido[4,5-d]pyrimid-ine-2,4(1H,3H)-dione (4d); and a pharmaceuticallyacceptable salt, ester, stereoisomer, or solvate thereof.

In an additional embodiment, the present subject matter relates to amethod of preparing a compound having the formula 4:

the method comprising:

-   mixing equimolar amounts of a substituted aromatic aldehyde having    the formula 1, a substituted barbituric acid derivative having the    formula 2, and a guanidine hydrochloride having the formula 3 to    obtain a first mixture;-   adding a mixture of water and ethanol along with a catalytic amount    of Ceric Ammonium Nitrate to the first mixture to obtain a reaction    mixture;-   stirring the reaction mixture until reaction completion; and-   filtering, washing with aqueous ethanol, and purifying the compound    of formula 4, wherein the reaction occurs according to the following    formula:

wherein:

-   R₁ is selected from the group consisting of hydrogen, hydroxy, and    benzyloxy;-   R₂ is selected from the group consisting of hydrogen, a C₁-C₆    straight or branched chain alkoxy, and benzyloxy;-   R₃ is selected from the group consisting of hydrogen and a C₁-C₆    straight or branched chain alkyl;-   R₄ is selected from the group consisting of hydrogen and a C₁-C₆    straight or branched chain alkyl; and-   Y is oxygen.

Further contemplated herein are pharmaceutical compositions containingthese compounds, as well as methods of treating an inflammatory disease,disorder, or condition by administering the present compounds to apatient in need thereof.

These and other features of the present subject matter will becomereadily apparent upon further review of the following specification.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

The following definitions are provided for the purpose of understandingthe present subject matter and for construing the appended patentclaims.

Definitions

Throughout the application, where compositions are described as having,including, or comprising specific components, or where processes aredescribed as having, including, or comprising specific process steps, itis contemplated that compositions of the present teachings can alsoconsist essentially of, or consist of, the recited components, and thatthe processes of the present teachings can also consist essentially of,or consist of, the recited process steps.

It is noted that, as used in this specification and the appended claims,the singular forms “a”, “an”, and “the” include plural references unlessthe context clearly dictates otherwise.

In the application, where an element or component is said to be includedin and/or selected from a list of recited elements or components, itshould be understood that the element or component can be any one of therecited elements or components, or the element or component can beselected from a group consisting of two or more of the recited elementsor components. Further, it should be understood that elements and/orfeatures of a composition or a method described herein can be combinedin a variety of ways without departing from the spirit and scope of thepresent teachings, whether explicit or implicit herein.

The use of the terms “include,” “includes”, “including,” “have,” “has,”or “having” should be generally understood as open-ended andnon-limiting unless specifically stated otherwise.

The use of the singular herein includes the plural (and vice versa)unless specifically stated otherwise. In addition, where the use of theterm “about” is before a quantitative value, the present teachings alsoinclude the specific quantitative value itself, unless specificallystated otherwise. As used herein, the term “about” refers to a ±10%variation from the nominal value unless otherwise indicated or inferred.

As used herein, “alkyl” refers to a straight-chain or branched saturatedhydrocarbon group. Examples of alkyl groups include methyl (Me), ethyl(Et), propyl (e.g., n-propyl and z′-propyl), butyl (e.g., n-butyl,z′-butyl, sec-butyl, tert-butyl), pentyl groups (e.g., n-pentyl,z′-pentyl, -pentyl), hexyl groups, and the like. In various embodiments,an alkyl group can have 1 to 40 carbon atoms (i.e., C₁-C₄₀ alkyl group),for example, 1-30 carbon atoms (i.e., C₁-C₃₀ alkyl group). In someembodiments, an alkyl group can have 1 to 6 carbon atoms, and can bereferred to as a “lower alkyl group” or a “C₁-C₆ alkyl group”. Examplesof lower alkyl groups include methyl, ethyl, propyl (e.g., n-propyl andz′-propyl), and butyl groups (e.g., n-butyl, z′-butyl, sec-butyl,tert-butyl). In some embodiments, alkyl groups can be substituted asdescribed herein. An alkyl group is generally not substituted withanother alkyl group, an alkenyl group, or an alkynyl group.

The term “substituted alkyl” as used herein refers to an alkyl group inwhich 1 or more (up to about 5, for example about 3) hydrogen atoms isreplaced by a substituent independently selected from the group: −O, −S,acyl, acyloxy, optionally substituted alkoxy, optionally substitutedamino (wherein the amino group may be a cyclic amine), azido, carboxyl,(optionally substituted alkoxy)carbonyl, amido, cyano, optionallysubstituted cycloalkyl, optionally substituted cycloalkenyl, halogen,hydroxyl, nitro, sulfamoyl, sulfanyl, sulfinyl, sulfonyl, and sulfonicacid. Some of the optional substituents for alkyl are hydroxy, halogenexemplified by chloro and bromo, acyl exemplified by methylcarbonyl;alkoxy, and heterocyclyl exemplified by morpholino and piperidino. Otheralkyl substituents as described herein may further be contemplated.

The term “optional” or “optionally” means that the subsequentlydescribed event or circumstance may or may not occur, and that thedescription includes instances where said event or circumstance occursand instances in which it does not. For example, “optionally substitutedalkyl” means either “alkyl” or “substituted alkyl,” as defined herein.

It will be understood by those skilled in the art with respect to anychemical group containing one or more substituents that such groups arenot intended to introduce any substitution or substitution patterns thatare sterically impractical and/or physically non-feasible.

The term “isomers” or “stereoisomers” as used herein relates tocompounds that have identical molecular formulae but that differ in thearrangement of their atoms in space. Stereoisomers that are not mirrorimages of one another are termed “diastereoisomers” and stereoisomersthat are non-superimposable mirror images are termed “enantiomers,” orsometimes optical isomers. A carbon atom bonded to four non-identicalsubstituents is termed a “chiral center.” Certain compounds herein haveone or more chiral centers and therefore may exist as either individualstereoisomers or as a mixture of stereoisomers. Configurations ofstereoisomers that owe their existence to hindered rotation about doublebonds are differentiated by their prefixes cis and trans (or Z and E),which indicate that the groups are on the same side (cis or Z) or onopposite sides (trans or E) of the double bond in the molecule accordingto the Cahn-Ingold-Prelog rules. All possible stereoisomers arecontemplated herein as individual stereoisomers or as a mixture ofstereoisomers.

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as commonly understood to one of ordinary skill inthe art to which the presently described subject matter pertains.

Where a range of values is provided, for example, concentration ranges,percentage ranges, or ratio ranges, it is understood that eachintervening value, to the tenth of the unit of the lower limit, unlessthe context clearly dictates otherwise, between the upper and lowerlimit of that range and any other stated or intervening value in thatstated range, is encompassed within the described subject matter. Theupper and lower limits of these smaller ranges may independently beincluded in the smaller ranges, and such embodiments are alsoencompassed within the described subject matter, subject to anyspecifically excluded limit in the stated range. Where the stated rangeincludes one or both of the limits, ranges excluding either or both ofthose included limits are also included in the described subject matter.

Throughout the application, descriptions of various embodiments use“comprising” language. However, it will be understood by one of skill inthe art, that in some specific instances, an embodiment canalternatively be described using the language “consisting essentiallyof” or “consisting of”.

“Subject” as used herein refers to any animal classified as a mammal,including humans, domestic and farm animals, and zoo, sports, and petcompanion animals such as household pets and other domesticated animalssuch as, but not limited to, cattle, sheep, ferrets, swine, horses,poultry, rabbits, goats, dogs, cats and the like.

“Patient” as used herein refers to a subject in need of treatment of acondition, disorder, or disease, such as an acute or chronic airwaydisorder or disease.

For purposes of better understanding the present teachings and in no waylimiting the scope of the teachings, unless otherwise indicated, allnumbers expressing quantities, percentages or proportions, and othernumerical values used in the specification and claims, are to beunderstood as being modified in all instances by the term “about”.Accordingly, unless indicated to the contrary, the numerical parametersset forth in the following specification and attached claims areapproximations that may vary depending upon the desired propertiessought to be obtained. At the very least, each numerical parametershould at least be construed in light of the number of reportedsignificant digits and by applying ordinary rounding techniques.

In an embodiment, the present subject matter relates to a compoundhaving the formula 4:

or a pharmaceutically acceptable salt, ester, stereoisomer, or solvatethereof, wherein:

-   R₁ is selected from the group consisting of hydrogen, hydroxy, and    benzyloxy;-   R₂ is selected from the group consisting of hydrogen, a C₁-C₆    straight or branched chain alkoxy, and benzyloxy;-   R₃ is selected from the group consisting of hydrogen and a C₁-C₆    straight or branched chain alkyl; and-   R₄ is selected from the group consisting of hydrogen and a C₁-C₆    straight or branched chain alkyl.

In one embodiment, R₂ can be selected from the group consisting ofhydrogen, methoxy, and benzyloxy. In a further embodiment, R₃ can beselected from the group consisting of hydrogen and methyl. In anadditional embodiment, R₄ can be selected from the group consisting ofhydrogen and methyl.

In certain embodiments, the compound can be selected from the groupconsisting of:5-(3-(Benzyloxy)phenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4a); 5-(4-(Benzyloxy)phenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione (4b);5-(3-Hydroxy-4-methoxyphenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4c);5-(3-Hydroxy-4-methoxyphenyl)-7-imino-1,3-dimethyl-7,8-dihydropyrimido[4,5-d]pyrimid-ine-2,4(1H,3H)-dione(4d); and a pharmaceutically acceptable salt, ester, stereoisomer, orsolvate thereof.

In another embodiment, the present subject matter relates to a compoundhaving the formula 4:

or a pharmaceutically acceptable salt, ester, stereoisomer, or solvatethereof, wherein:

-   R₁ is selected from the group consisting of hydrogen, hydroxy, and    benzyloxy;-   R₂ is selected from the group consisting of hydrogen, methoxy, and    benzyloxy;-   R₃ is selected from the group consisting of hydrogen and methyl; and-   R₄ is selected from the group consisting of hydrogen and methyl.

In a further embodiment, the present subject matter relates to acompound selected from the group consisting of:5-(3-(Benzyloxy)phenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4a);5-(4-(Benzyloxy)phenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4b);5-(3-Hydroxy-4-methoxyphenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4c);5-(3-Hydroxy-4-methoxyphenyl)-7-imino-1,3-dimethyl-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4d); and a pharmaceutically acceptable salt, ester, stereoisomer, orsolvate thereof.

Said differently, the present subject matter can relate to compounds offormula 4, i.e.,5-(substitutedphenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dioneanalogues, selected from the group consisting of:

and a pharmaceutically acceptable salt, ester, stereoisomer, or solvatethereof.

It is to be understood that the present subject matter covers allcombinations of substituent groups referred to herein.

The present compounds may contain, e.g., when isolated in crystallineform, varying amounts of solvents. Accordingly, the present subjectmatter includes all solvates of the present compounds of formula 4 andpharmaceutically acceptable stereoisomers, esters, and/or salts thereof.Hydrates are one example of such solvates.

Further, the present subject matter includes all mixtures of possiblestereoisomers of the embodied compounds, independent of the ratio,including the racemates.

Salts of the present compounds, or salts of the stereoisomers thereof,include all inorganic and organic acid addition salts and salts withbases, especially all pharmaceutically acceptable inorganic and organicacid addition salts and salts with bases, particularly allpharmaceutically acceptable inorganic and organic acid addition saltsand salts with bases customarily used in pharmacy.

Examples of acid addition salts include, but are not limited to,hydrochlorides, hydrobromides, phosphates, nitrates, sulfates, acetates,trifluoroacetates, citrates, D-gluconates, benzoates,2-(4-hydroxy-benzoyl)benzoates, butyrates, subsalicylates, maleates,laurates, malates, lactates, fumarates, succinates, oxalates, tartrates,stearates, benzenesulfonates (besilates), toluenesulfonates (tosilates),methanesulfonates (mesilates) and 3-hydroxy-2-naphthoates.

Examples of salts with bases include, but are not limited to, lithium,sodium, potassium, calcium, aluminum, magnesium, titanium, ammonium,meglumine and guanidinium salts. The salts include water-insoluble and,particularly, water-soluble salts.

The present compounds, the salts, the stereoisomers and the salts of thestereoisomers thereof may contain, e.g., when isolated in crystallineform, varying amounts of solvents. Included within the present scopeare, therefore, all solvates of the compounds of formula 4, as well asthe solvates of the salts, the stereoisomers and the salts of thestereoisomers of the compounds of formula 4.

The present compounds may be isolated and purified in a manner known perse, e.g., by distilling off the solvent in vacuo and recrystallizing theresidue obtained from a suitable solvent or subjecting it to one of thecustomary purification methods, such as column chromatography on asuitable support material.

Salts of the compounds of formula 4 and the stereoisomers thereof can beobtained by dissolving the free compound in a suitable solvent (by wayof non-limiting example, a ketone such as acetone, methylethylketone ormethylisobutylketone; an ether such as diethyl ether, tetrahydrofuraneor dioxane; a chlorinated hydrocarbon such as methylene chloride orchloroform; a low molecular weight aliphatic alcohol such as methanol,ethanol or isopropanol; a low molecular weight aliphatic ester such asethyl acetate or isopropyl acetate; or water) which contains the desiredacid or base, or to which the desired acid or base is then added. Theacid or base can be employed in salt preparation, depending on whether amono- or polybasic acid or base is concerned and depending on which saltis desired, in an equimolar quantitative ratio or one differingtherefrom. The salts are obtained by filtering, reprecipitating,precipitating with a non-solvent for the salt or by evaporating thesolvent. Salts obtained can be converted into the free compounds which,in turn, can be converted into salts. In this manner, pharmaceuticallyunacceptable salts, which can be obtained, for example, as processproducts in the manufacturing on an industrial scale, can be convertedinto pharmaceutically acceptable salts by processes known to the personskilled in the art.

Pure diastereomers and pure enantiomers of the present compounds can beobtained, e.g., by asymmetric synthesis, by using chiral startingcompounds in synthesis and by splitting up enantiomeric anddiastereomeric mixtures obtained in synthesis. Preferably, the purediastereomeric and pure enantiomeric compounds are obtained by usingchiral starting compounds in synthesis.

Enantiomeric and diastereomeric mixtures can be split up into the pureenantiomers and pure diastereomers by methods known to a person skilledin the art. Preferably, diastereomeric mixtures are separated bycrystallization, in particular fractional crystallization, orchromatography. Enantiomeric mixtures can be separated, e.g., by formingdiastereomers with a chiral auxiliary agent, resolving the diastereomersobtained and removing the chiral auxiliary agent. As chiral auxiliaryagents, for example, chiral acids can be used to separate enantiomericbases and chiral bases can be used to separate enantiomeric acids viaformation of diastereomeric salts. Furthermore, diastereomericderivatives such as diastereomeric esters can be formed fromenantiomeric mixtures of alcohols or enantiomeric mixtures of acids,respectively, using chiral acids or chiral alcohols, respectively, aschiral auxiliary agents. Additionally, diastereomeric complexes ordiastereomeric clathrates may be used for separating enantiomericmixtures. Alternatively, enantiomeric mixtures can be split up usingchiral separating columns in chromatography. Another suitable method forthe isolation of enantiomers is enzymatic separation.

The present compounds can show promising anti-inflammatory activitybetween millimolar to micromolar concentrations compared to standardanti-inflammatory drugs. Some of the selected lead compounds can besuccessfully taken forward to develop novel anti-inflammatory drugcandidates.

In one embodiment, the present subject matter relates to a method ofpreparing a compound having the formula 4:

the method comprising:

-   mixing equimolar amounts of a substituted aromatic aldehyde having    the formula 1, a substituted barbituric acid derivative having the    formula 2, and a guanidine hydrochloride having the formula 3 to    obtain a first mixture;-   adding a mixture of water and ethanol along with a catalytic amount    of Ceric Ammonium Nitrate to the first mixture to obtain a reaction    mixture;-   stirring the reaction mixture until reaction completion; and-   filtering, washing with aqueous ethanol, and purifying the compound    of formula 4, wherein the reaction occurs according to the following    formula:

wherein:

-   R₁ is selected from the group consisting of hydrogen, hydroxy, and    benzyloxy;-   R₂ is selected from the group consisting of hydrogen, a C₁-C₆    straight or branched chain alkoxy, and benzyloxy;-   R₃ is selected from the group consisting of hydrogen and a C₁-C₆    straight or branched chain alkyl;-   R₄ is selected from the group consisting of hydrogen and a C₁-C₆    straight or branched chain alkyl; and-   Y is oxygen.

In this regard, the mixture of water and ethanol can have a ratio of8:2, v/v. In another embodiment, the reaction mixture can be stirred atroom temperature for at least about 4 hours. In a further embodiment,the present methods can obtain the compound of formula 4 at a yield ofabout 78% to about 89%.

In another embodiment, the present subject matter is directed topharmaceutical compositions comprising a therapeutically effectiveamount of the compounds as described herein together with one or morepharmaceutically acceptable carriers, excipients, or vehicles. In someembodiments, the present compositions can be used for combinationtherapy, where other therapeutic and/or prophylactic ingredients can beincluded therein.

The present subject matter further relates to a pharmaceuticalcomposition, which comprises at least one of the present compoundstogether with at least one pharmaceutically acceptable auxiliary.

In an embodiment, the pharmaceutical composition comprises one or two ofthe present compounds, or one of the present compounds.

Non-limiting examples of suitable excipients, carriers, or vehiclesuseful herein include liquids such as water, saline, glycerol,polyethyleneglycol, hyaluronic acid, ethanol, and the like. Suitableexcipients for nonliquid formulations are also known to those of skillin the art. A thorough discussion of pharmaceutically acceptableexcipients and salts useful herein is available in Remington'sPharmaceutical Sciences, 18th Edition. Easton, Pa., Mack PublishingCompany, 1990, the entire contents of which are incorporated byreference herein.

The present compounds are typically administered at a therapeutically orpharmaceutically effective dosage, e.g., a dosage sufficient to providetreatment for cancer. Administration of the compounds or pharmaceuticalcompositions thereof can be by any method that delivers the compoundssystemically and/or locally. These methods include oral routes,parenteral routes, intraduodenal routes, and the like.

While human dosage levels have yet to be optimized for the presentcompounds, generally, a daily dose is from about 0.01 to 10.0 mg/kg ofbody weight, for example about 0.1 to 5.0 mg/kg of body weight. Theprecise effective amount will vary from subject to subject and willdepend upon the species, age, the subject's size and health, the natureand extent of the condition being treated, recommendations of thetreating physician, and the therapeutics or combination of therapeuticsselected for administration. The subject may be administered as manydoses as is required to reduce and/or alleviate the signs, symptoms, orcauses of the disease or disorder in question, or bring about any otherdesired alteration of a biological system.

In employing the present compounds for treatment of cancer, anypharmaceutically acceptable mode of administration can be used withother pharmaceutically acceptable excipients, including solid,semi-solid, liquid or aerosol dosage forms, such as, for example,tablets, capsules, powders, liquids, suspensions, suppositories,aerosols or the like. The present compounds can also be administered insustained or controlled release dosage forms, including depotinjections, osmotic pumps, pills, transdermal (includingelectrotransport) patches, and the like, for the prolongedadministration of the compound at a predetermined rate, preferably inunit dosage forms suitable for single administration of precise dosages.

The present compounds may also be administered as compositions preparedas foods for foods or animals, including medical foods, functional food,special nutrition foods and dietary supplements. A “medical food” is aproduct prescribed by a physician that is intended for the specificdietary management of a disorder or health condition for whichdistinctive nutritional requirements exist and may include formulationsfed through a feeding tube (referred to as enteral administration orgavage administration).

A “dietary supplement” shall mean a product that is intended tosupplement the human diet and may be provided in the form of a pill,capsule, tablet, or like formulation. By way of non-limiting example, adietary supplement may include one or more of the following dietaryingredients: vitamins, minerals, herbs, botanicals, amino acids, anddietary substances intended to supplement the diet by increasing totaldietary intake, or a concentrate, metabolite, constituent, extract, orcombinations of these ingredients, not intended as a conventional foodor as the sole item of a meal or diet. Dietary supplements may also beincorporated into foodstuffs, such as functional foods designed topromote control of glucose levels. A “functional food” is an ordinaryfood that has one or more components or ingredients incorporated into itto give a specific medical or physiological benefit, other than a purelynutritional effect. “Special nutrition food” means ingredients designedfor a particular diet related to conditions or to support treatment ofnutritional deficiencies.

Generally, depending on the intended mode of administration, thepharmaceutically acceptable composition will contain about 0.1% to 90%,for example about 0.5% to 50%, by weight of a compound or salt of thepresent compounds, the remainder being suitable pharmaceuticalexcipients, carriers, etc.

One manner of administration for the conditions detailed above is oral,using a convenient daily dosage regimen which can be adjusted accordingto the degree of affliction. For such oral administration, apharmaceutically acceptable, non-toxic composition is formed by theincorporation of any of the normally employed excipients, such as, forexample, mannitol, lactose, starch, magnesium stearate, sodiumsaccharine, talcum, cellulose, sodium crosscarmellose, glucose, gelatin,sucrose, magnesium carbonate, and the like. Such compositions take theform of solutions, suspensions, tablets, dispersible tablets, pills,capsules, powders, sustained release formulations and the like.

The present compositions may take the form of a pill or tablet and thusthe composition may contain, along with the active ingredient, a diluentsuch as lactose, sucrose, dicalcium phosphate, or the like; a lubricantsuch as magnesium stearate or the like; and a binder such as starch, gumacacia, polyvinylpyrrolidine, gelatin, cellulose and derivativesthereof, and the like.

Liquid pharmaceutically administrable compositions can, for example, beprepared by dissolving, dispersing, etc. an active compound as definedabove and optional pharmaceutical adjuvants in a carrier, such as, forexample, water, saline, aqueous dextrose, glycerol, glycols, ethanol,and the like, to thereby form a solution or suspension. If desired, thepharmaceutical composition to be administered may also contain minoramounts of nontoxic auxiliary substances such as wetting agents,emulsifying agents, or solubilizing agents, pH buffering agents and thelike, for example, sodium acetate, sodium citrate, cyclodextrinederivatives, sorbitan monolaurate, triethanolamine acetate,triethanolamine oleate, etc.

For oral administration, a pharmaceutically acceptable non-toxiccomposition may be formed by the incorporation of any normally employedexcipients, such as, for example, pharmaceutical grades of mannitol,lactose, starch, magnesium stearate, talcum, cellulose derivatives,sodium crosscarmellose, glucose, sucrose, magnesium carbonate, sodiumsaccharin, talcum and the like. Such compositions take the form ofsolutions, suspensions, tablets, capsules, powders, sustained releaseformulations and the like.

For a solid dosage form, a solution or suspension in, for example,propylene carbonate, vegetable oils or triglycerides, may beencapsulated in a gelatin capsule. Such diester solutions, and thepreparation and encapsulation thereof, are disclosed in U.S. Pat. Nos.4,328,245; 4,409,239; and 4,410,545, the contents of each of which areincorporated herein by reference. For a liquid dosage form, thesolution, e.g., in a polyethylene glycol, may be diluted with asufficient quantity of a pharmaceutically acceptable liquid carrier,e.g., water, to be easily measured for administration.

Alternatively, liquid or semi-solid oral formulations may be prepared bydissolving or dispersing the active compound or salt in vegetable oils,glycols, triglycerides, propylene glycol esters (e.g., propylenecarbonate) and the like, and encapsulating these solutions orsuspensions in hard or soft gelatin capsule shells.

Other useful formulations include those set forth in U.S. Pat. Nos. Re.28,819 and 4,358,603, the contents of each of which are herebyincorporated by reference.

Another manner of administration is parenteral administration, generallycharacterized by injection, either subcutaneously, intramuscularly orintravenously. Injectables can be prepared in conventional forms, eitheras liquid solutions or suspensions, solid forms suitable for solution orsuspension in liquid prior to injection, or as emulsions. Suitableexcipients are, for example, water, saline, dextrose, glycerol, ethanolor the like. In addition, if desired, the pharmaceutical compositions tobe administered may also contain minor amounts of non-toxic auxiliarysubstances such as wetting or emulsifying agents, pH buffering agents,solubility enhancers, and the like, such as for example, sodium acetate,sorbitan monolaurate, triethanolamine oleate, cyclodextrins, etc.

Another approach for parenteral administration employs the implantationof a slow-release or sustained-release system, such that a constantlevel of dosage is maintained. The percentage of active compoundcontained in such parenteral compositions is highly dependent on thespecific nature thereof, as well as the activity of the compound and theneeds of the subject. However, percentages of active ingredient of 0.01%to 10% in solution are employable and will be higher if the compositionis a solid which will be subsequently diluted to the above percentages.The composition may comprise 0.2% to 2% of the active agent in solution.

Nasal solutions of the active compound alone or in combination withother pharmaceutically acceptable excipients can also be administered.

Formulations of the active compound or a salt may also be administeredto the respiratory tract as an aerosol or solution for a nebulizer, oras a microfine powder for insufflation, alone or in combination with aninert carrier such as lactose. In such a case, the particles of theformulation have diameters of less than 50 microns, for example lessthan 10 microns.

The present compounds have valuable pharmaceutical properties, whichmake them commercially utilizable. Accordingly, the present subjectmatter further relates to use of the present compounds for the treatmentof diseases such as inflammatory diseases, disorders, or conditions.

An embodiment of the present subject matter is directed to a method oftreating an inflammatory disease, disorder, or condition in a patient,comprising administering to a patient in need thereof a therapeuticallyeffective amount of a compound as described herein. A therapeuticallyeffective amount of the compound or pharmaceutically acceptablecomposition or an amount effective to treat a disease, such as aninflammatory disease, disorder, or condition, may be determinedinitially from the Examples described herein and adjusted for specifictargeted diseases using routine methods.

In an embodiment, the inflammatory disease, disorder, or condition canbe pain. In another embodiment, the inflammatory disease, disorder, orcondition can be an acute or chronic inflammation.

The present subject matter further relates to a method of treating orpreventing a disease comprising administering to a patient in needthereof a therapeutically effective amount of at least one of thecompounds herein.

In particular, the present subject matter relates to a method oftreating one of the above-mentioned diseases or disorders comprisingadministering to a patient in need thereof a therapeutically effectiveamount of at least one of the compounds herein.

In the above methods, the patient is preferably a mammal, morepreferably a human. Furthermore, in the above methods, at least one ofthe present compounds can be used. In an embodiment, one or two of thepresent compounds are used, or one of the present compounds is used.Similarly, one or more of the present compounds can be used incombination therapy with one or more additional active agents.

The present subject matter can be better understood by referring to thefollowing examples.

EXAMPLES Example 1

Preparation of5-(substitutedphenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dioneanalogues (4a-d)

Compounds 4a-4d were synthesized according to the following Scheme 1.

General synthetic procedure for the synthesis of5-(substitutedphenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dioneanalogues (4a-d)

A mixture of equimolar amounts of substituted aromatic aldehyde (1, 1equiv.), substituted barbituric acid derivative (2, 1 equiv.), andguanidine hydrochloride (3, 1 equiv.) was taken in a round-bottom flask,and a mixture of water and ethanol (8:2, v/v) was added along with acatalytic amount of Ceric Ammonium Nitrate. The reaction medium wasstirred at room temperature for 4 h. Completion of the reaction wasmonitored by TLC (EtOAc: hexane, 2:8), and the final product wasfiltered, washed with aqueous ethanol, and used for purification.

The characterization details of compounds 4a-4d are reported below.

5-(3-(Benzyloxy)phenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4a). Yellow solid; Yield: 78%; Melting point: 206-208° C.; FT-IR (KBrcm⁻¹): 3234 (−NH), 3109 (Ar−H), 2934 (C−H), 1689 (C=O), 1677 (C=N); ¹HNMR (DMSO-d₆, 400 MHz): δ=11.38 (s, 1H, −NH), 11.23 (s, 1H, −NH), 8.21(s, 1H, −NH), 7.88 (s, 1H, −NH), 7.59 (d, 1H, Ar−H, J=8.0 Hz), 7.43-7.28(m, 7H, Ar−H), 7.16 (dd, 1H, Ar−H, J=8.0 and 2.4 Hz), 5.09 (s, 2H,−OCH₂); ¹³C NMR (DMSO-d₆, 100 MHz): δ=193.5, 163.9, 162.2, 158.3, 155.0,150.7, 137.3, 134.4, 131.0, 129.7, 129.0, 128.5, 126.8, 123.3, 119.9,119.6, 119.2, 114.4, 69.9; LCMS (m/z): 361.80 [M]⁺; Elemental analysisof C₁₉H₁₅N₅O₃, Calcd: C: 63.15; H: 4.18; N: 19.38. Found: C: 63.18; H:4.15; N: 19.39.

5-(4-(Benzyloxy)phenyl)-7-imino-7,8-dihydropyrimido[4,5]dipyrimidine-2,4(1H,3H)-dione(4b). Yellow solid; Yield: 89%; Melting point: 138-140° C.; FT-IR (KBrcm⁻¹): 3184 (−NH), 3064 (Ar−H), 2902 (C−H), 1686 (C=O), 1631 (C=N); ¹HNMR (DMSO-d₆, 400 MHz): δ 11.33 (s, 1H, −NH), 11.20 (s, 1H, −NH), 8.36(d, 2H, J=8.8 Hz), 8.25 (s, 1H, −NH), 7.88 (s, 1H, −NH), 7.47-7.35 (m,5H, Ar−H), 7.13 (d, 2H, Ar−H, J=8.8 Hz), 5.23 (s, 2H, −OCH₂); ¹³C NMR(DMSO-d₆, 100 MHz): δ=164.4, 163.0, 162.6, 155.4, 150.7, 137.9, 136.8,129.0, 128.6, 128.4, 125.8, 116.1, 115.1, 70.1; LCMS (m/z): 362.42[M+H]⁺; Elemental analysis of C₁₉H₁₅N₅O₃, Calcd: C: 63.15; H: 4.18; N:19.38. Found: C: 63.14; H: 4.17; N: 19.37.

5-(3-Hydroxy-4-methoxyphenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4c). Yellow solid; Yield: 83%; Melting point: 268-270° C.; FT-IR (KBrcm⁻¹): 3518 (−OH), 3216 (−NH), 3116 (Ar−H), 1689 (C=O), 1588 (C=N); ¹HNMR (DMSO-d₆, 400 MHz): δ=11.24 (s, 1H, −NH), 11.12 (s, 1H, −NH), 9.42(s, 1H, −NH), 8.11 (s, 1H, Ar−H), 8.08 (d, 1H, Ar−H, J=1.6 Hz), 7.68(dd, 2H, methoxyphenyl-H, J =8.8 and 1.8 Hz), 7.02 (d, 1H,methoxyphenyl-H, J=8.4 Hz), 3.84 (s, 3H, −OCH₃); ¹³C NMR (DMSO-d₆, 100MHz): δ=164.6, 162.7, 156.1, 153.4, 150.8, 146.3, 130.9, 125.9, 120.9,115.6, 111.8, 56.3; LCMS (m/z): 301.13 [M]⁺; Elemental analysis ofC₁₃H₁₁N₅O₄, Calcd: C: 51.83; H: 3.68; N: 23.25. Found: C: 51.84; H:3.66; N: 23.27.

5-(3-Hydroxy-4-methoxyphenyl)-7-imino-1,3-dimethyl-7,8-dihydropyrimido[4,5-d]pyrimid-ine-2,4(1H,3H)-dione(4d). Yellow solid; Yield: 89%; Melting point: 188-190° C.; FT-IR (KBrcm ⁻¹): 3522 (−OH), 3248 (−NH), 3126 (Ar−H), 1718 (C=O), 1595 (C=N); ¹HNMR (DMSO-d₆, 400 MHz): δ=9.43 (s, 1H, −OH), 8.18 (s, 2H, −NH), 8.06 (d,1H, methoxyphenyl-H, J=2.0 Hz), 7.66 (dd, 1H, methoxyphenyl-H, J=8.8 and2.4 Hz), 7.02 (d, 1H, methoxyphenyl-H, J=8.8 Hz), 3.84 (s, 3H, −OCH₃),3.17 (s, 6H, −NCH₃); ¹³C NMR (DMSO-d₆, 100 MHz): δ=163.2, 161.3, 156.9,153.5, 151.7, 146.3, 130.9, 125.9, 120.7, 115.3, 111.8, 56.3, 29.2,28.6; LCMS (m/z): 330.38 [M+H]⁺; Elemental analysis of C₁₅H₁₅N₅O₄,Calcd: C: 54.71; H: 4.59; N: 21.27. Found: C: 54.74; H: 4.56; N: 21.26.

Example 2 Anti-inflammatory Activity

The anti-inflammatory activity IC₅₀ values for the designed compounds4a, 4b, 4c, and 4d were evaluated in μM, as summarized below in Table 1.

TABLE 1 Compound code Compound Structure IC₅₀ (μM) 4a

0.2483 4b

0.0909 4c

0.2184 4d

0.1059

It is to be understood that the present compounds, compositions, andmethods are not limited to the specific embodiments described above, butencompasses any and all embodiments within the scope of the genericlanguage of the following claims enabled by the embodiments describedherein, or otherwise shown in the drawings or described above in termssufficient to enable one of ordinary skill in the art to make and usethe claimed subject matter.

We claim:
 1. A compound having the formula 4:

or a pharmaceutically acceptable salt, ester, stereoisomer, or solvatethereof, wherein: R₁ is selected from the group consisting of hydrogen,hydroxy, and benzyloxy; R₂ is selected from the group consisting ofhydrogen, a C₁-C₆ straight or branched chain alkoxy, and benzyloxy; R₃is selected from the group consisting of hydrogen and a C₁-C₆ straightor branched chain alkyl; and R₄ is selected from the group consisting ofhydrogen and a C₁-C₆ straight or branched chain alkyl.
 2. The compoundof claim 1, wherein R₂ is selected from the group consisting ofhydrogen, methoxy, and benzyloxy.
 3. The compound of claim 1, wherein R₃is selected from the group consisting of hydrogen and methyl.
 4. Thecompound of claim 1, wherein R₄ is selected from the group consisting ofhydrogen and methyl.
 5. The compound of claim 1, wherein the compound isselected from the group consisting of:5-(3-(Benzyloxy)Phenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4a);5(4-(Benzyloxy)phenyl)-7-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4b);5-(3-Hydroxy-4-methoxyphenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4c);5-(3-Hydroxy-4-methoxyphenyl)-7-imino-1,3-dimethyl-7,8-dihydropyrimido[4,5-d]pyrimid-ine-2,4(1H,3H)-dione(4d); and a pharmaceutically acceptable salt, ester, stereoisomer, orsolvate thereof.
 6. A pharmaceutically acceptable composition comprisinga therapeutically effective amount of the compound of claim 1 and apharmaceutically acceptable carrier.
 7. A compound having the formula 4:

or a pharmaceutically acceptable salt, ester, stereoisomer, or solvatethereof, wherein: R₁ is selected from the group consisting of hydrogen,hydroxy, and benzyloxy; R₂ is selected from the group consisting ofhydrogen, methoxy, and benzyloxy; R₃ is selected from the groupconsisting of hydrogen and methyl; and R₄ is selected from the groupconsisting of hydrogen and methyl.
 8. The compound of claim 7, whereinthe compound is selected from the group consisting of:5-(3-(Benzyloxy)Phenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4a);5-(4-(Benzyloxy)phenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4b);5-(3-Hydroxy-4-methoxyphenyl)-7-imino-7,8-dihydropyrimido[4,5-d]pyrimidine-2,4(1H,3H)-dione(4c);5-(3-Hydroxy-4-methoxyphenyl)-7-imino-1,3-dimethyl-7,8-dihydropyrimido[4,5-d]pyrimid-ine-2,4(1H,3H)-dione(4d); and a pharmaceutically acceptable salt, ester, stereoisomer, orsolvate thereof.
 9. A pharmaceutically acceptable composition comprisinga therapeutically effective amount of the compound of claim 7 and apharmaceutically acceptable carrier.
 10. A method of preparing acompound having the formula 4:

the method comprising: mixing equimolar amounts of a substitutedaromatic aldehyde having the formula 1, a substituted barbituric acidderivative having the formula 2, and a guanidine hydrochloride havingthe formula 3 to obtain a first mixture; adding a mixture of water andethanol along with a catalytic amount of Ceric Ammonium Nitrate to thefirst mixture to obtain a reaction mixture; stirring the reactionmixture until reaction completion; and filtering, washing with aqueousethanol, and purifying the compound of formula 4, wherein the reactionoccurs according to the following formula:

wherein: R₁ is selected from the group consisting of hydrogen, hydroxy,and benzyloxy; R₂ is selected from the group consisting of hydrogen, aC₁-C₆ straight or branched chain alkoxy, and benzyloxy; R₃ is selectedfrom the group consisting of hydrogen and a C₁-C₆ straight or branchedchain alkyl; R₄ is selected from the group consisting of hydrogen and aC₁-C₆ straight or branched chain alkyl; and Y is oxygen.
 11. The methodof claim 10, wherein the mixture of water and ethanol has a ratio of8:2, v/v.
 12. The method of claim 10, wherein the reaction mixture isstirred at room temperature for at least about 4 hours.
 13. The methodof claim 10, wherein: R₁ is selected from the group consisting ofhydrogen, hydroxy, and benzyloxy; R₂ is selected from the groupconsisting of hydrogen, methoxy, and benzyloxy; R₃ is selected from thegroup consisting of hydrogen and methyl; and R₄ is selected from thegroup consisting of hydrogen and methyl.
 14. The method of claim 10,wherein the method obtains the compound of formula 4 at a yield of about78% to about 89%.